Study of the mechanism of hemostatic effect of desmopressin.

Intravenous injection of 1 microg/kg desmopressin to rabbits not only accelerated clotting of arterial blood (maximally by 59.16 +/- 8.53% after 1 h), but also increased the number of microvesicles containing the integral enzyme 5'-nucleotidase in the arterial blood from the initial level of 36.26 +/- 8.08 ncat/liter to a maximum of 99.65 +/- 15.8 ncat/liter after 15 min.

Cell microvesicles during experimental endotoxemia.

The dynamics of microvesicle formation in arterial blood in generalized Schwartzman phenomenon was studied. Successive (with 24-h interval) intravenous injections of endotoxin to rabbits in a dose of 1 mg/kg and 3 mg/kg caused an increase in the content of microvesicles in the blood, some of them containing ecto-5'-nucleotidase. Biphasic changes in arterial blood clotting time and erythrocyte hemolysis were observed.

Effect of lupus anticoagulant on the interaction of prothrombin with platelets.

We studied binding of (125)I-labeled prothrombin to platelets in the presence of circulating endogenous lupus anticoagulant. Lupus anticoagulant modulated the interaction and increased affinity of prothrombin for calcium-independent binding sites on platelets. The number of these sites decreased, while the total number of calcium-dependent binding sites increased. Our results indicate that lupus anticoagulant plays a role in the binding of prothrombin to nonactivated platelets.

[Interaction of human factor X with thromboplastin]. / Vzaimodeistvie faktora X cheloveka s troboplastinom.

The binding of 125I-labeled human factor X to native and papaine-treated tissue tromboplastin in the presence of CaCl2 or EDTA was studied. The Scatchard analysis suggests the existence of high (Kd=l,8 x10(-9) M) and low affinity binding sites on the thromboplastin surface. The removal of Ca2+ reduced affinity of factor X to the high affinity sites. This was accompanied by some increase of their number. Proteolysis by papaine decreased affinity of high affinity sites and caused the increase of their number in the presence of Ca2+. In the absence of Ca2+ the affinity remained unchanged, but the number of sites decreased. At low concentrations of factor X positive cooperativity for high affinity binding sites was observed. It did not depend on the presence of Ca2+. The results indirectly confirm the role of hydrophobic interactons in Ca2+ dependent binding of factor X to thromboplastin and the fact that heterogeneity of this binding is determined by mesophase structure of the thromboplastin phospholipids.

Effect of radiation on prothrombin interactions with cell membrane fragments.

Binding of (125)I-labeled prothrombin to normal and radiation-damaged (1 and 3.5 Gy) fragments of cell membranes was studied. Irradiation significantly increased adsorption capacity of cell membranes for (125)I-prothrombin. It was concluded that phase rearrangement of phospholipids serves as the molecular basis for increased thrombogenicity of cell membranes.

[Effect of Glycine max lectin on the interaction of prothrombin and erythrocytes]. / Vliianie lektina Clycine max na vzaimodeistvie protrombina s éritrotsitami.

Native porcine erythrocytes do not initiate blood coagulation, though even the weak association (Kd = 4,25 +/- 9,35 microM) of prothrombin with their surface which is limited to the projection of two phospholipid polar head groups onto the external cell membrane, exerts a slight but authentic influence on the acceleration of blood clotting. There are 11.9 x 10(7) sites for 125I-prothrombin on one erythrocytes. At physiological concentration of prothrombin the surface of erythrocytes is far from saturation. The binding of 125I-labelled prothrombin to erythrocytes after their surface was covered by lectin Glycine max by means of the accessible residues of N-acetyl-D-galactosamine and D-galactose, glycoproteins and dlycolipids can never become saturated with any used concentration of the ligand. In the presence of the suspension of erythrocytes treated lectin platelet free plasma coagulates at the same speed as the control plasma.

[The rate of physiological fibrinolysis in the mesenteric and renal vessels of rabbits]. / Skorost' fiziologicheskogo fibrinoliza v mezenterial'nykh i pochechnykh sosudakh u krolikov.

In the rabbit vessels of a. mesenterica, total lysis of fibrin occurred within 60-90 minutes whereas in the renovascular basin the process took 30 minutes longer. This seems to be due to synthesis of a plasminogen activator.

[Interaction of prothrombin with erythrocytes and alveolar macrophages]. / Vzaimodeistvie protrombina s éritrotsitami i al'veoliarnymi makrofagami.

A study was made of porcine 125I-prothrombin interaction with red cells and alveolar macrophages in the presence of CaCl2 or EDTA. Prothrombin binding with red cells was similar to that to alveolar macrophages (Kd = 3.7 x 10(-6) M and 1.9 x 10(-6) M, respectively). These parameters did not depend on the presence of Ca2+ ions in the medium. Red cell hemolysis led to diminished affinity of prothrombin to blood ghost surfaces.

[Interaction of human thrombin I fragment, prethrombin I, and alpha-thrombin with tissue thromboplastin]. / Vzaimodeistvie fragmenta I protrombina, pretrombina I, and alpha- trombina cheloveka s tkanevym tromboplastinom.

The binding of 125I-labeled prothrombin fragment I. prethrombin I and alpha-thrombin to native and papain-treated tissue thromboplastin in the presence of CaCl2 of EDTA was studied. The experimental curves plotted in the Scatchard coordinates testify to the presence in thromboplastin of two types of fragment I binding sites: those with a high (Kd = 7.6 x 10(-6) M) and moderate (Kd = 1.3 x 10(-8) M) binding affinity. The parameters of fragment I binding and their changes reproduced, for the most part, the mode of prothrombin binding observed in previous studies. The experimental results provide indirect evidence in favour of a hydrophobic role of Ca(2+)-dependent binding of prothrombin fragment I to thromboplastin. The binding of prethrombin I was nonspecific and Ca(2+)-independent, whereas alpha-thrombin showed a relatively high level of nonspecific electrostatic binding which was competitively inhibited by Ca2+. Thromboplastin proteins interacted (both directly and in a Ca(2+)-independent fashion) with all the prothrombin derivatives under study.

[Irreversible pH-denaturation of tissue thromboplastin]. / Neobratimaia pH-denaturatsiia tkanevogo tromboplastina.

Irreversible denaturation of tissue thromboplastin from human brain occurred at pH values below 5.0 and above 11.0. Coagulating activity of total phospholipid fraction of thromboplastin was decreased after incubation in acid or alkaline media.

[Functional concept of the initiation of blood coagulation by cell membranes]. / Funktsional'naia kontseptsiia initsirovaniia svertyvaniia krovi kletochnymi membranami.

According to the conception proposed, blood coagulation is initiated by Ca(2+)-induced rearrangement of the bilayer structure of native cell membranes into heterophasic. The thrombogenic action is realized through Ca2+ entering the cell cytoplasm. This leads to phosphatidyl serine translocation into the surface monolayer of the membrane, its clustering and the arising of phosphatidyl ethanolamine mesophases. Vitamin K-dependent coagulation factors are bound at the boundaries of clusters and mesophases of these phospholipids through Ca2+ ions. They form enzyme complexes functioning due to the matrix structure of phospholipid surface as units of thrombin generation.

[The determination of tissue thromboplastin activity in cells]. / Opredelenie aktivnosti tkanevogo tromboplastina v kletkakh.

A method for estimation of thromboplastin activity in blood and tissue cells has been developed, experimentally and theoretically substantiated. Blood plasma free from blood coagulation contact phase factors is used as a substrate for the detection of thromboplastic activities of blood and tissue cells.

[Interaction of lectins with tissue thromboplastin]. / Vzaimodeistvie lektinov s tkanevym tromboplastinom.

Exo- and endolectins affined to N-acetylgalactosamine, N-acetylglucosamine, galactose, mannose and fucose, added in vitro to suspension of tissue thromboplastin from human brain, suppress its blood coagulation activity by 50-88% (p less than 0.05). This result has been considered as an argument for the advantage of the opinion that the specific blood coagulation centre of apoprotein III is located at the extracellular side of the cytoplasmic membrane.

[Interaction of human prothrombin with tissue thromboplastin]. / Vzaimodeistvie protrombina cheloveka s tkanevym tromboplastinom.

The binding of 125I-labeled human prothrombin to native and papain-treated tissue thromboplastin in the presence of CaCl2 or EDTA was studied. The Scatchard plots for the protein binding suggest the presence at thromboplastin surface of two types of binding sites, high affinity [Kd(app) = 7.4.10(-8) M] and moderate affinity [Kd(app) = 7.9.10(-5) M]. The removal of Ca2+ did not influence the Kd (values for these) sites but markedly reduced their number. Proteolysis by papain caused a decrease in the affinity of high affinity sites without affecting the Kd values of the moderate affinity sites yet caused a proportional increase in the number of both high and moderate affinity sites in the presence of Ca2+. At low prothrombin concentrations a positive cooperativity of protein binding at high affinity sites in the presence of Ca2+ was observed.

[Participation of alveolar macrophages in the catabolism of fibrinogen and fibrin]. / Uchastie al'veoliarnykh makrofagov v katabolizme fibrinogena i fibrina.

The penetration of FITC-fibrinogen in rabbit alveolar macrophages depends on Ca2+ and Mg2+. Therefore there is reason to believe the process is not restricted only by diffusion on concentration gradient, but pinocytosis takes place, which depends Ca2+. The absorption of FITC-label from fibrin by alveolar macrophages is more slow than from fibrinogen. Soluble high-molecular products of fibrin degradation can be trapped by pinocytosis. On the basis of such studies it was assumed that degradation of fibrin proceeds parietally.

[The expression and degradation of tissue thromboplastin]. / Ekspressiia i degradatsiia tkanevogo tromboplastina.

The highest disintegration rate of FITC-thromboplastin occurred in the organs rich in RES-cells. The half-disintegration period was found to be 58 +/- 12 min in liver, 98 +/- 48 min in spleen, 262 +/- 36 min in kidneys, 310 +/- 114 min in lung microcirculatory flow. Several pathways were found of the thromboplastin tissue local disintegration mediated via proteolytic and lipolytic enzymes released from endothelial cells and leucocytes: endocytosis of RES-cells followed by intracellular cleavage by lysosomal enzymes; and gradual blood transfer of thromboplastin particles from the organs with moderate disintegration rate to the organs capable of rapid disintegration.

[Effect of fibronectin on fibrinogen conversion into fibrin]. / Vliianie fibronektina na prevrashchenie fibrinogena v fibrin.

The effects of fibronectin on fibrinogen clotting induced by thrombin or reptilase and on fibrin monomer polymerization in a pure system in the absence of factor XIIIa were studied. It was shown that within a broad range of concentrations and molar ratios of the mixed proteins, fibronectin does not alter significantly the fibrinogen clotting time either under thrombin or under reptilase action. The effect of fibronectin on the fibrin self-assembly consists in a slight acceleration of this process, whose degree is directly dependent on the fibronectin/fibrin monomer molar ratio as well as on the absolute fibrin monomer content at a constant molar ratio. The stimulating effect of fibronectin is amplified by Ca2+. The experimental results suggest that fibronectin can noncovalently bind the fibrin monomer and/or intermediate polymers in the non-enzymatic phase of fibrinogen conversion to fibrin.

[Complex-formation with hyaluronic acid as a cause of fibronectin heterogeneity in the synovial fluid]. / Kompleksoobrazovanie s gialuronovoi kislotoi kak prichina geterogennosti fibronektina v sinovial'noi zhidkosti.

Heterogeneity of synovial fluid fibronectin was studied by means of Laurell cross-immunoelectrophoresis in patients with rheumatoid arthritis, posttraumatic synovitis and other arthropathies. Prior to hyaluronidase treatment all the synovial fluid samples exhibited the fibronectin heterogeneity, which disappeared after the action of hyaluronidase. The data obtained suggest that complexes of fibronectin and hyaluronic acid are responsible for physico-chemical heterogeneity of fibronectin in synovial fluid.